Replication - Part 1 Flashcards
1
Q
Prokaryotic genome organization
A
- Lower DNA content
- Main genome: single major DNA molecule (cccDNA)
- Operons
- Few repetitive sequences: no introns
- Other genes: plasmid
2
Q
Eukaryotic genome organization
A
- Higher DNA content
- Enclosed in a nuclear membrane
- Several linear DNA, chromosome
- Organization of coding sequences
- With telomere
3
Q
DNA replication is __
A
Semi-conservative
4
Q
What is maintained inthe DNA
A
Specific base pairing
5
Q
Needed for initiation of replication
A
Primer
6
Q
Direction of newly synthesized DNA strand
A
5’-3’ direction
7
Q
Replication of DNA is __
A
Semi-discontinuous
8
Q
Grows continuously
A
Leading strand
9
Q
Grows in segments
A
Lagging strand
10
Q
Connects the okazaki fragments
A
DNA ligase
11
Q
How are nucleotides become linked
A
Condensation reaction to form phosphodiester linkage
12
Q
Where replication begins and proceeds
A
- begins at an origin
- proceeds bidirectionally
13
Q
What is involved in DNA replication
A
Several enzymes and proteins (replisomes)
14
Q
How are bacterial genes usually written
A
3 lowercase italicized letters
15
Q
Example of how the gene and gene products are written
A
dnaA (gene) -> Dna A (protein)
16
Q
Replication proteins
A
Replisome
17
Q
What are the replisomes
A
- Initiation protein
- DNA helicase/ DNA unwinding proteins
- Helix destabilizing proteins or single-strand binding proteins (SSBP)
- Primase
- DNA polymerases
- DNA ligase
- Topoisomerases
18
Q
Initiation protein of E.coli
A
dnaA protein
19
Q
- recognizes the OriC
- binds to it (~20-30 dnaA)
A
dnaA protein
20
Q
- local denaturation of DNA helix
- Requires ATP
- Binds ssDNA (not dsDNA)
A
DNA helicases/DNA unwinding proteins
21
Q
2 helicases of E.coli
A
- Helicase II
- Rep protein
22
Q
- attaches to template for lagging strand
- moves 5’–>3’
A
Helicase II
23
Q
- attached to template for leading strand
- moves 3’–>5’
A
Rep protein
24
Q
Bind to sites where template has been unwound but not yet duplicated
A
Helix Destabilizing Proteins or Single-strand Binding Proteins (SSBP)
25
Functions of Helix Destabilizing Proteins or Single-strand Binding Proteins (SSBP)
1. prevent reannealing and tangling of ssDNA
2. prevent formation of 2º structure
3. protect ssDNA from nuclease degradation
26
Major SSB of eukaryotes
Replication protein A (RPA)
27
SSBPs are required to __ the unwound DNA
Iron out
28
Generate primers
Primases
29
- short RNA chain (4-12 nts)
- provides the 3’OH end needed by DNA polymerase during elongation
Primer
30
- monomeric or heteromultimeric
- 3 enzyme activities
DNA polymerase
31
3 enzymatic activities of DNA Polymerase
1. 5'-->3' polymerization activity
2. 3'-->5' exonuclease activity
3. 5'-->3' exonuclease activity
32
Add nucleotides onto the 3'OH end of a primer --> synthesize new DNA strands
5'-->3' polymerization activity
33
property of DNA pol to remain attached to the template and incorporate nucleotides before detaching
Processivity
34
1 mistake per 10^9 or 10^10 nucs added
Fidelity
35
Proofreading
3'-->5' exonuclease activity
36
- removes RNA primers at the 5’ end
- not present in eukaryotic DNA polymerase
5’-->3’ exonuclease activity
37
enzyme that perform 5'-3' exonuclease activity in eukaryotes
1. Rnase H
2. Flap endonuclease 1 (FEN1)
3. DNA pol delta
38
- remove RNA primers
- detects RNA-DNA hybrid in eukaryotes
Rnase H
39
removes remaining bits or RNA primer in eukaryotes
Flap endonuclease 1 (FEN1)
40
add/fill in the gaps from removal of primer in eukaryotes
DNA pol delta
41
3 main DNA polymerases for replication
1. DNA pol I
2. DNA pol II
3. DNA pol III
42
Major repair enzyme
DNA pol I, II
43
Elongating enzyme
DNA pol III
44
Gene of DNA pol I
_polA_
45
Gene of DNA pol II
_polB_
46
Gene of DNA pol III
_polC_
47
SOS repair
DNA pol IV, V
48
Gene of DNA pol IV
_dinB_
49
Gene of DNA pol V
umuD'2C
50
- Replicates both strands simultaneously
- A dimer of 10 proteins
Bacterial DNA Polymerase III Holoenzyme
51
Core enzyme of Bacterial DNA Polymerase III Holoenzyme
- α – (5’->3' polymerization)
ε - (3’->5' exonuclease/proofreading)
- θ stimulates ε
52
Sliding clamp of Bacterial DNA Polymerase III Holoenzyme
β2
53
purpose of sliding clamp β2
helps the holoenzyme
54
Clamp loader of Bacterial DNA Polymerase III Holoenzyme
(δδ'χΨ)2τ2γ2
55
Keeps DNA polymerase attached to the template to allow duplication of long stretches of DNA
Sliding clamp
56
Assembles the sliding clamp to the DNA
Clamp loader
57
E. coli sliding clamp = __
eukaryotic PCNA
58
Separates large fragment and 5'->3' exonuclease
Subtilisin (proteolysis)
59
Used in in vitro synthesis of dsDNA from ssDNA
Klenow fragment
60
Eukaryotic DNA polymerases
α, β, γ, δ, ε, etc
(10 DNA polymerases)
61
Location of DNA pol α
Nuclear
62
Location of DNA pol β
Nuclear
63
Location of DNA pol γ
Mitochondrial
64
Location of DNA pol δ
Nuclear
65
Location of DNA pol ε
Nuclear
66
function of DNA pol α
priming and replication of lagging strand
67
function of DNA pol β
repair
68
function of DNA pol γ
mitochondrial
69
function of DNA pol δ
replication of leading strand
70
function of DNA pol ε
probably replication
71
relative activity of α
~80%
72
relative activity of DNA pol β
10-15%
73
relative activity of DNA pol γ
2-15%
74
MW of α
300,000
75
MW of β
40,000
76
MW of γ
180-300,000
77
MW of δ
170-230,000
78
MW of ε
255,000
79
No. of subunits of α
4 (several)
80
No. of subunits of β, γ, δ
1
81
DNA polymerases with 3'-5' exonuclease activity
γ, δ, ε
82
seals nicks using ATP or NAD
DNA ligase
83
what does the DNA ligase use to seal nicks
ATP or NAD
84
- relaxes DNA
- removes negative supercoils
topoisomerases
85
topoisomerase that relax only one strand
topoisomerase I
86
topoisomerase that relax both strands
topoisomerase II
87
in _E.coli_, topoisomerase or gyrase is involved in what?
deconcatenation
88
process of releasing the final products of circular DNA replication
deconcatenation
89
what stage of the cell cycle does DNA replication happen
S phase
90
stages of replication
1. initiation
2. elongation
3. termination
91
recognition of positions on a DNA molecule where replication will begin
initiation
92
copying of parent polynucleotide (template) at the replication fork
elongation
93
completion
termination
